Optimising vitrification of human oocytes using multiple cryoprotectants and morphological and functional assessment

Oocyte vitrification is a clinical practice that allows preservation of fertility potential in women. Vitrification involves quick cooling using high concentrations of cryoprotectants to minimise freezing injuries. However, high concentrations of cryoprotectants have detrimental effects on oocyte quality and eventually the offspring. In addition, current assessment of oocyte quality after vitrification is commonly based only on the morphological appearance of the oocyte, raising concerns regarding its efficiency. Using both morphological and functional assessments, the present study investigated whether combinations of cryoprotectants at lower individual concentrations result in better cryosurvival rates than single cryoprotectants at higher concentrations. Surplus oocytes from IVF patients were vitrified within 24h after retrieval using the Cryotop method with several cryoprotectants, either individually or in combination. The morphological and functional quality of the vitrified oocytes was investigated using light microscopy and computer-based quantification of mitochondrial integrity, respectively. Oocyte quality was significantly higher using a combination of cryoprotectants than vitrification with individual cryoprotectants. In addition, the quality of vitrified oocyte varied depending on the cryoprotectants and type of combination used. The results of the present study indicate that observations based purely on the morphological appearance of the oocyte to assess the cryosurvival rate are insufficient and sometimes misleading. The outcome will have a significant implication in the area of human oocyte cryopreservation as an important approach for fertility preservation.